Following elucidation of the regulation of the lactose operon in Escherichia coli, studies on the metabolism of many sugars were initiated in the early 1960s. The catabolic pathways of D-gluconate and of the two hexuronates, D-glucuronate and D-galacturonate, were investigated. The post genomic era has renewed interest in the study of these sugar acids and allowed the complete characterization of the D-gluconate pathway and the discovery of the catabolic pathways for L-idonate, D-glucarate, galactarate, and ketogluconates. Among the various sugar acids that are utilized as sole carbon and energy sources to support growth of E. coli, galacturonate, glucuronate, and gluconate were shown to play an important role in the colonization of the mammalian large intestine. In the case of sugar acid degradation, the regulators often mediate negative control and are inactivated by interaction with a specific inducer, which is either the substrate or an intermediate of the catabolism. These regulators coordinate the synthesis of all the proteins involved in the same pathway and, in some cases, exert crosspathway control between related catabolic pathways. This is particularly well illustrated in the case of hexuronide and hexuronate catabolism. The structural genes encoding the different steps of hexuronate catabolism were identified by analysis of numerous mutants affected for growth with galacturonate or glucuronate. E. coli is able to use the diacid sugars D-glucarate and galactarate (an achiral compound) as sole carbon source for growth. Pyruvate and 2-phosphoglycerate are the final products of the D-glucarate/galactarate catabolism.

Environmental citrate or malonate is degraded by a variety of aerobic or anaerobic bacteria. For selected examples, the genes encoding the specific enzymes of the degradation pathway are described together with the encoded proteins and their catalytic mechanisms. Aerobic bacteria degrade citrate readily by the basic enzyme equipment of the cell if a specific transporter for citrate is available. Anaerobic degradation of citrate in Klebsiella pneumoniae requires the so-called substrate activation module to convert citrate into its thioester with the phosphoribosyl dephospho-CoA prosthetic group of citrate lyase. The citryl thioester is subsequently cleaved into oxaloacetate and the acetyl thioester, from which a new citryl thioester is formed as the turnover continues. The degradation of malonate likewise includes a substrate activation module with a phosphoribosyl dephospho-CoA prosthetic group. The machinery gets ready for turnover after forming the acetyl thioester with the prosthetic group. The acetyl residue is then exchanged by a malonyl residue, which is easily decarboxylated with the regeneration of the acetyl thioester. This equipment suffices for aerobic growth on malonate, since ATP is produced via the oxidation of acetate. Anaerobic growth on citrate or malonate, however, depends on additional enzymes of a so-called energy conservation module. This allows the conversion of decarboxylation energy into an electrochemical gradient of Na+ ions. In citrate-fermenting K. pneumoniae, the Na+ gradient is formed by the oxaloacetate decarboxylase and mainly used to drive the active transport of citrate into the cell. To use this energy source for this purpose is possible, since ATP is generated by substrate phosphorylation in the well-known sequence from pyruvate to acetate. In the malonate-fermenting bacterium Malonomonas rubra, however, no reactions for substrate level phosphorylation are available and the Na+ gradient formed in the malonate decarboxylation reaction must therefore be used as the driving force for ATP synthesis.

During fermentative growth, Escherichia coli degrades carbohydrates via the glycolytic route into two pyruvate molecules. Pyruvate can be reduced to lactate or nonoxidatively cleaved by pyruvate formate lyase into acetyl-coenzyme A (acetyl-CoA) and formate. Acetyl-CoA can be utilized for energy conservation in the phosphotransacetylase (PTA) and acetate kinase (ACK) reaction sequence or can serve as an acceptor for reducing equivalents gathered during pyruvate formation, through the action of alcohol dehydrogenase (AdhE). Formic acid is strongly acidic and has a redox potential of −420 mV under standard conditions and therefore can be classified as a high-energy compound. Its disproportionation into CO2 and molecular hydrogen (Em,7 −420 mV) via the formate hydrogenlyase (FHL) system is therefore of high selective value. The FHL reaction involves the participation of at least seven proteins, most of which are metalloenzymes, with requirements for iron, molybdenum, nickel, or selenium. Complex auxiliary systems incorporate these metals. Reutilization of the hydrogen evolved required the evolution of H2 oxidation systems, which couple the oxidation process to an appropriate energy-conserving terminal reductase. E. coli has two hydrogen-oxidizing enzyme systems. Finally, fermentation is the "last resort" of energy metabolism, since it gives the minimal energy yield when compared with respiratory processes. Consequently, fermentation is used only when external electron acceptors are absent. This has necessitated the establishment of regulatory cascades, which ensure that the metabolic capability is appropriately adjusted to the physiological condition. Here we review the genetics, biochemistry, and regulation of hydrogen metabolism and its hydrogenase maturation system.

Glutamate, aspartate, asparagine, L-alanine, and D-alanine are derived from intermediates of central metabolism, mostly the citric acid cycle, in one or two steps. While the pathways are short, the importance and complexity of the functions of these amino acids befit their proximity to central metabolism. Inorganic nitrogen (ammonia) is assimilated into glutamate, which is the major intracellular nitrogen donor. Glutamate is a precursor for arginine, glutamine, proline, and the polyamines. Glutamate degradation is also important for survival in acidic environments, and changes in glutamate concentration accompany changes in osmolarity. Aspartate is a precursor for asparagine, isoleucine, methionine, lysine, threonine, pyrimidines, NAD, and pantothenate; a nitrogen donor for arginine and purine synthesis; and an important metabolic effector controlling the interconversion of C3 and C4 intermediates and the activity of the DcuS-DcuR two-component system. Finally, L- and D-alanine are components of the peptide of peptidoglycan, and L-alanine is an effector of the leucine responsive regulatory protein and an inhibitor of glutamine synthetase (GS). This review summarizes the genes and enzymes of glutamate, aspartate, asparagine, L-alanine, and D-alanine synthesis and the regulators and environmental factors that control the expression of these genes. Glutamate dehydrogenase (GDH) deficient strains of E. coli, K. aerogenes, and S. enterica serovar Typhimurium grow normally in glucose containing (energy-rich) minimal medium but are at a competitive disadvantage in energy limited medium. Glutamate, aspartate, asparagine, L-alanine, and D-alanine have multiple transport systems.

This review focuses on recent data concerning the ecological factors that determine the distribution of Escherichia coli and the genetic structures of naturally occurring E. coli populations. It summarizes some of the older literature concerning the dynamics of E. coli populations within a host and poses some questions that arise from our more recently acquired understanding of the factors affecting the genetic structures of E. coli populations. Multilocus enzyme electrophoresis (MLEE) studies indicate that E. coli, relative to other members of the family Enterobacteriaceae, exhibits a moderate degree of genetic diversity. The existence of subspecific structure in E. coli has for the most part been determined by largely neutral in its effects on the fitness of a strain. The consequences for E. coli of the transition between its primary and secondary habitats are of considerable practical significance for water quality assessment and disease transmission. E. coli causes a significant fraction of human bacterial disease and is responsible for two main types of disease in humans and domestic animals: diarrheal disease and extraintestinal infections. The observed distribution of strains from the different E. coli genetic groups indicates that they have different life history tactics and ecological niches. A and B1 strains appear to be generalists, as they can be recovered from any vertebrate group. Group B2 and D strains appear to be more specialized, as they are largely restricted to endothermic vertebrates.

Microbes produce an extraordinary array of microbial defense systems. These include broad-spectrum classical antibiotics critical to human health concerns; metabolic by-products, such as the lactic acids produced by lactobacilli; lytic agents, such as lysozymes found in many foods; and numerous types of protein exotoxins and bacteriocins. The abundance and diversity of this biological arsenal are clear. Lactic acid production is a defining trait of lactic acid bacteria. Bacteriocins are found in almost every bacterial species examined to date, and within a species, tens or even hundreds of different kinds of bacteriocins are produced. Halobacteria universally produce their own version of bacteriocins, the halocins. Streptomycetes commonly produce broad-spectrum antibiotics. It is clear that microbes invest considerable energy in the production and elaboration of antimicrobial mechanisms. What is less clear is how such diversity arose and what roles these biological weapons play in microbial communities. One family of microbial defense systems, the bacteriocins, has served as a model for exploring evolutionary and ecological questions. In this review, current knowledge of how the extraordinary range of bacteriocin diversity arose and is maintained in one species of bacteria, Escherichia coli, is assessed and the role these toxins play in mediating microbial dynamics is discussed.

In this chapter we review the literature with respect to what is known about how Escherichia coli colonizesthe mammalian intestine. We begin with a brief discussion of the mammalian large intestine, the major site that commensal strains of E. coli colonize. Next, evidence is discussed showing that, in order to colonize, E. coli must be able to penetrate and grow in the mucus layer of the large intestine. This is followed by discussions of colonization resistance, i.e., factors that are involved in the ability of a complete microbiota (microflora) to resist colonization by an invading bacterium, the advantages and disadvantages of the in vivo colonization models used in colonization research, the initiation and maintenance stages of E. coli colonization, and the rate of E. coli growth in the intestine. The next two sections of the chapter discuss the role of motility in colonization and how adhesion to mucosal receptors aids or inhibits penetration of the intestinal mucus layer and thereby either promotes or prevents E. coli colonization. Finally, the contribution of nutrition to the ability of E. coli to colonize is discussed based on the surprising finding that different nutrients are used by E. coli MG1655, a commensal strain, and by E. coli EDL933, an enterohemorrhagic strain, to colonize the intestine.

During the early evolution of life, gene duplication, the production of two copies of a DNA sequence, allowed the rapid diversification of enzymatically catalyzed reactions and an increase in genome size, providing also material for the invention of new enzymatic properties and complex regulatory and developmental patterns. A duplication may involve (i) a part of a gene, (ii) a whole gene, (iii) DNA stretches including two or more genes involved in the same or in different metabolic pathways, (iv) entire operons, (v) a part of a chromosome, (vi) an entire chromosome, and finally (vii) the whole genome. Therefore, any DNA sequence may undergo a duplication event(s), but the fate of the replicate depends on whether it provides an evolutionary advantage to the host cell. Two hypotheses on the origin and evolution of metabolic pathways exist. The first one, the Horowitz retrograde hypothesis, predicts that an entire metabolic route was assembled by successive duplications of an ancestral gene in a backward fashion, starting with the synthesis of the final product, then the penultimate pathway intermediate, and so on down the pathway to the initial precursor. The patchwork hypothesis is based on the duplication(s) of ancestral gene(s) leading to the progressive increasing of specificity of low-specific enzymes, which then may be recruited to catalyze similar reactions in different metabolic pathways or sequential steps in the same route.

Enzymatic inactivation of antibiotics occurs with several of the natural product antibiotic classes but has not yet been observed as a major route of resistance development for the classes of synthetic antibacterials: the sulfamethoxazole-trimethoprim combination, the fluoroquinolones, or the oxazolidinones. The most widespread mode of clinical resistance development to β-lactam antibiotics is the expression of β-lactamases that hydrolyze the antibiotic. Two approaches have been taken in the decades since lactam-resistant clinical isolates began to diminish the efficacy of penicillins and cephalosporins as antibiotics. The first has been to develop semisynthetic β-lactams which were slower substrates for attack by the hydrolytic lactamases. The second approach has been to screen for inhibitors and inactivators of lactamase activity and then combine these molecules with a β-lactam. β-Lactamase genes can be embedded in bacterial chromosomes, such as the ampC gene in enteric bacteria or the blaZ gene in Staphylococcus aureus, or they can be carried on multiple-copy plasmids or transposons, as is the case for the TEM-1 bla gene in a variety of high-level penicillin-resistant gram-negative bacteria found in clinical isolates. In Escherichia coli the ampG, ampD, and ampR genes control expression of the ampC-encoding β-lactamase. In S. pneumoniae external penicillin leads to an increase in autolytic peptidoglycan hydrolase activity and subsequent vulnerability to osmotic lysis and death. Three kinds of enzymatic modifications of OH and NH2 groups on aminoglycosides are common determinants of resistance and represent variants of normal electrophilic group transfer enzymes that participate in primary metabolism.

Legionellae were initially characterized by criteria typically used to describe an autonomous unicellular organism. Bacteria of the genus Legionella were described as gram-negative, aerobic, and rod shaped with one or more polar or lateral flagella. A more complete ecological profile of legionellae will provide the scientific basis for selecting the most appropriate host for pathogenesis studies as well as studies to identify procedures to control amplification of the bacteria in certain environments. Analysis of bacterial group behavior represents the most recent facet in the study of the ecology of legionellae. In building water systems, legionellae are most frequently detected in biofilms of plumbing fixtures and heating, ventilating, and air-conditioning (HVAC) equipment. The complex nutrients available with biofilms have led some researchers to propose that the biofilm might allow the survival and multiplication of legionellae outside a host cell. If legionellae can multiply extracellularly within biofilms, the study and characterization of this phenomenon could have tremendous impact on control strategies for the prevention of legionellosis. Researchers have only begun to characterize the interaction of legionellae with the microbial community. Controlling legionellae within biofilms may lead to the most effective control measures to prevent legionellosis.

This chapter aims at determining the susceptibility of biofilm-associated Legionella pneumophila to free chlorine and monochloramine. A biofilm reactor was developed to grow biofilms containing L. pneumophila on 24 replicate stainless-steel surfaces in potable water. The ability of this reactor to produce reproducible biofilms was validated by the fact that the standard deviations of the base biofilm densities on stainless-steel coupons (n = 3) ranged from 0.06 to 0.18 log CFU per coupon. When Legionella containing biofilms were exposed to the same dosages of monochloramine for identical contact periods, the treatments were significantly more effective. In summary, the authors have shown that biofilm-associated L. pneumophila are significantly less susceptible to chlorine than are planktonic L. pneumophila, while susceptibility of planktonic and biofilm-associated L. pneumophila to monochloramine are similar. When monochloramine and free chlorine were compared under identical conditions, monochloramine was significantly more effective, indicating that monochloramine may be an effective disinfectant for the inactivation of L. pneumophila within potable water distribution systems. Further research using open system biofilm reactors and model distribution systems is needed to determine the utility of monochloramine as a disinfectant against biofilm-associated L. pneumophila.

The immunogenicity of self-like microbial molecules is strikingly illustrated in several cases of murine and human systemic lupus erythematosus (SLE). Several indirect arguments support the idea that microbial agents influence the course of antiphospholipid syndrome (APS). An association between APS and pathogens was documented, such as hepatitis C virus, Salmonella lipopolysaccharide, and Mycoplasma penetrans, a rare bacterium that has so far only been found in human immunodeficiency virus (HlV)-infected persons and that was isolated from the blood and throat of a non-HIV-infected patient with primary APS. The molecular basis of antigen mimicry by anti-idiotypic antibodies was studied extensively. On the basis of Jerne's theory, after immunization with an autoantibody that carries a specific idiotype (Abl), naive mice develop an antiautoantibody (anti-Id; which is also known as Ab2) and then generate anti-anti-Id (Ab3) a few weeks later. Immunization of naive mice with anticardiolipin β2GPI-dependent MAbs and polyclonal antibodies or their corresponding scFv, such as Abl, resulted in the production in the inoculated mice of autoantibody directed to cardiolipin and to the cardiolipin β2GPI-dependent antibody Ab3. Studies on experimental lupus and experimental APS prove the existence of molecular mimicry between pathogens and the autoantigens involved in experimental lupus and APS. Recognition of multiple antigens and epitopes is evident in insulin-dependent diabetes mellitus, SLE, APS, rheumatoid arthritis, PBC, and probably most autoimmune diseases, with spreading of the named epitope leading to autoantibody spread.

Dramatic increases in cps expression correlate with increased capsule synthesis and generally can be ascribed to either of two control points for the regulatory system. Capsule synthesis under all these conditions is completely dependent on RcsB; RcsA appears to act as an accessory factor that allows modulation of RcsB activity. When high-level expression of cps-lac fusions is selected at elevated temperatures, the primary location of the resulting mutations is in rcsC, the gene immediately clockwise to rcsB. Overproduction of RcsF increases capsule synthesis twofold, and mutations in rcsF decrease capsule synthesis two to threefold. Although there is no evidence for phosphorylation of RcsA, the temperature sensitivity of capsule synthesis is apparently best explained by temperature sensitivity of RcsA activity. The RcsB protein is relatively abundant in cells, and it is unclear whether there is any significant regulation of its activity during various growth conditions. RcsA is normally synthesized in very low amounts, and the protein is difficult to detect in wild-type cells, due to both low levels of synthesis and rapid degradation.